The incidence of bloodstream infections caused by Candida species has increased dramatically; these organisms now account for 10% of all bloodstream isolates. The mortality associated with candidemia approaches 40%, even with therapy. Because of this unacceptably high mortality, new strategies to treat and prevent this infection are necessary. Our goal is to identify new Candida albicans virulence genes and to determine the mechanisms by which they contribute to pathogenicity. This information holds promise to identify new targets for antifungal strategies. Our hypothesis is that the ability of C. albicans to invade and damage host cells is critical for the organism to establish and maintain a deep-seated infection. Support for this hypothesis comes from our finding that many C. albicans mutants with decreased virulence in the mouse model of hematogenously disseminated infection also have reduced ability to invade and damage endothelial cells in vitro. Furthermore, we have screened a collection of random C. albicans homozygous insertion mutants to identify strains with impaired capacity to damage endothelial cells in vitro. We discovered that zed1/zed1 and cka2/cka2 insertion mutants caused much less endothelial cell damage than did the isogenic control strain. Also, the zed1/zed1 insertion mutant had significantly attenuated virulence in mice (the virulence of the cka2/cka2 strain has not yet been tested). These results strongly suggest that C. albicans genes required for in vitro endothelial cell damage are also required for virulence. Our objective is to use in vitro studies of the interactions between C. albicans and endothelial cells to define mechanisms of host-pathogen interaction. In this project, we will 1) identify the endothelial cell receptors that C. albicans uses to invade this host cell; 2) determine the mechanisms by which C. albicans Zed1p, Cka2p, and other newly identified gene products contribute to virulence; 3) use defined C. albicans mutants to elucidate functional relationships among C. albicans virulence regulators, endothelial cell receptors, and endothelial cell damage; and 4) use a random insertional mutagenesis approach to identify new C. albicans genes that promote endothelial cell damage in vitro and virulence.